Approximately 154 years ago, we initiated the first clinical trail investigating allogeneic stem cell transplantation in patients with treatment refractory metastatic renal cell carcinoma (RCC). Definitive evidence for an allogeneic graft-versus-RCC effect has been demonstrated with complete regression of large metastasis observed in some patients. We have subsequently conducted studies investigating the immunological mechanisms accounting for tumor regression in patients demonstrating an anti-RCC effect in attempts to identify both the effector cell populations mediating these regressions as well as their target antigens. We demonstrated that cytotoxic T-cell clones with specificity for minor antigens are capable of killing RCC cells in patients having a GVT effect post transplant. Using c-DNA expression cloning, we have recently identified a novel tumor antigen derived from a human endogenous retrovirus over-expressed in RCC cells called HERV-E CT-RCC. This antigen is not expressed on normal tissues and therefore could potentially serve as a target for a future kidney cancer vaccine. Work to identify other antigens derived from this HERV-E that are expressed on common HLA molecules as well as efforts to develop a monoclonal antibody that recognizes HERV-E derived proteins that are selectively expressed in RCC are ongoing. We have also sought to characterize the mechanisms accounting for selective expression of the CT-RCC HERV-E in RCC. Recently, we found that inactivation of the VHL tumor suppressor gene, over-expression of the hypoxia-inducible transcription factor (HIF) HIF-2, and demethylation of the HERV-E 5LTR results in the selective expression of HERV-E in clear cell carcinoma. These studies have provided valuable insights needed for ongoing translational research aimed at boosting human immunity against antigenic components of this provirus. We are also exploring methods to sensitize solid tumors to NK cell attack by altering the phenotype of tumor cells through targeted gene induction. Recently we showed that bortezomib and depsipeptide sensitize tumors to NK cell cytotoxity by enhancing NK-cell mediated TRAIL killing. This sensitization appears to overcome NK inhibition that is mediated through KIR-KIR ligand interactions. We have also developed a method to expand by > 4 logs NK cells from healthy donors for adoptive infusion in future NK-cell based adoptive immunotherapy trials. We now have developed a method to expand human NK cells by 100 to 1000 fold initially in bags and now in G-REX flasks using GMP conditions. A phase I study entitled Safety and the anti- tumor effects of escalating doses of adoptively infused ex vivo expanded autologous natural killer cells against metastatic cancers or hematological malignancies sensitized to NK TRAIL cytotoxicity with Bortezomib was initiated in 2008 and is currently accruing patients. With the exception of thyroiditis, infusions of up to 2.5x108 expanded NK cells/kg have been well tolerated and provided early evidence for mediating anti-tumor effects in patients with cancer. We have also initiated NK cell studies in rhesus macaques to evaluate methods to optimize the antitumor function and in vivo persistence and tumor homing of adoptively infused autologous NK cells. We have also recently partnered with Dr. Ola Landgren from the NCI to explore KIR blockade with the monoclonal antibody IPH 2101 (Innate Pharma) as a method to enhance autologous NK cell killing of smoldering myeloma. Finally, our lab has recently shown that mRNA electroporation of ex vivo expanded NK cells using the clinical-grade Maxcyte transfection system is highly efficient at transfecting NK cells opening numerous new possibilities to genetically reprogram NK cells to advance the field of NK cell-based cancer immunotherapy. My group has continued to investigate methods to improve allogeneic HCT for heavily pretreated patients with bone marrow failure syndromes (BMFS) including paroxysmal nocturnal hemoglobinuria (PNH) and ATG-refractory severe aplastic anemia (SAA). In a cohort of 56 heavily transfused and HLA alloimmunized patients with BMFS undergoing a novel fludarabine-based HCT, graft rejection did not occur and 87% of the patients survived disease free with a median follow-up of 4.5 years. Despite this excellent survival, approximately 70% of patients developed chronic GVHD. In a multivariate analysis, we found that rapid donor T-cell engraftment (d 95% donor T-cell chimerism by day 30) was associated with the development of chronic GVHD (adjusted HR 3.03, p=0.04). To test whether delaying the speed of donor T-cell engraftment will prevent chronic GVHD, we have initiated a new clinical trial for BMFS patients which uses a modified donor allograft containing high doses of G-CSF mobilized CD34+ selected cells combined with a one log reduced dose of non-G-CSF mobilized T-cells. Preliminary data suggest this transplant approach slows the speed of donor T-cell engraftment compared to a T-cell replete trransplant, significantly reducing the risk of both acute and chronic GVHD Transplantation using unrelated cord blood (UCB) has been shown to be a reasonable alternative transplant approach for patients with a variety of hematological malignancies and non-malignant hematological disorders who lack an HLA matched related or unrelated stem cell donor. Recent data show leukemia-free survival in adult patients with AML and ALL is comparable to that of recipients receiving fully matched unrelated donor transplants. However, for patients with SAA undergoing UCB transplantation, results have been disappointing. Investigators from the EBMT and Eurocord have reported engraftment rates of only 51% and survival rates of only 38% in SAA recipients of single or dual CB transplants. In order to harness the advantage of UCB availability, to overcome the disadvantage of delayed neutrophil recovery of UCB transplantation, and to provide a back up stem cell source should the CB unit fail to engraft, we have recently initiated a clinical trial that evaluates the co-administration of unrelated umbilical cord blood and a relatively low number of highly purified haploidentical peripheral blood CD34+ cells from a related donor as a method to promote rapid neutrophil recovery. Subjects receive a novel non-myeloablative immunosuppressive conditioning regimen of cyclophosphamide, fludarabine, horse ATG and one dose of total body irradiation (200cGy) followed by an infusion of the allografts. The haploidentical stem cell product is T-cell depleted and enriched for CD34+ cells using the Miltenyi CliniMacs system. To reduce TRM secondary to prolonged neutropenia associated with conventional UCB transplantation, haploidentical CD34+ stem cells are co-infused with a single UCB unit (serologically matched at d 4/6 HLA loci). The primary endpoint will be donor engraftment by day 42. This study is now accruing patients with 20/20 intial patients treated having donor engraftment by day 22 and with a median followup of 2.3 years, survival has been excellent at 87%.